Clinically significant alcoholic (A) liver damage (LD), secondary to a hepatic necroinflammatory cascade (HNIC), occurs only in a subset of alcoholics. Hence, factors other than ethanol (E) must be involved. Hypothesis: The key cofactor for ALD is a breakdown of gut barrier integrity ("leaky gut") due to chronic E use, which allows intestinal endotoxin to reach the liver &initiate a HNIC;this leakiness is due to cytoskeletal instability caused by oxidation of cytoskeletal proteins which is elicited by E-induced gut iNOS upregulation &nitric oxide (NO) overproduction. We found: 1} in man, gut leakiness in alcoholics with LD but not in those without LD or in nonalcoholics with LD;2} in rats, E-induced leaky gut is associated with LD;reversal of gut leakiness attenuates LD;3} in intestinal monolayers, E-induced iNOS upregulation causes cytoskeletal &barrier disruption. We will continue to use this successful translational approach (monolayers, rats &man) to test our current hypotheses. Aims: (1) To see if, in a larger sample, a leaky gut: a) occurs only in alcoholics with LD &precedes cirrhosis b) persists during abstinence &after liver transplant for ALD, c) correlates quantitatively with LD severity, d) is associated with NO overproduction &HNIC, e) is more pronounced in females. We predict that gut leakiness (excess urinary lactulose, mannitol &sucralose levels after oral sugar load): i) is seen only in alcoholics with LD, precedes cirrhosis;ii) correlates with severity of LD (clinical parameters, liver enzymes);iii) is associated with NO overproduction (gut mucosal NO), serum endotoxin &HNIC (high neopterin/cytokines). (2) To see if, in rats, prevention of E-induced leaky gut also prevents E-induced LD &if a hyperactive, NO pathway is involved. We predict that in E-fed rats with LD: i) leaky gut, endotoxemia, HNIC, upregulation of intestinal iNOS, NO overproduction &oxidation of actin &tubulin occurs;ii) preventing gut leakiness (by oats, iNOS inhibitors or Arginine) prevents LD. (3) To see, using monolayers of wild type ((inhibitors) &transfected cells, if E-induced iNOS upregulation &its consequences (assessed by cytoskeletal oxidation/disarray &barrier disruption) are mediated by NF-kappaB activation. We predict i) E activates NF-kappaB by degrading IkappaBalpha;ii) preventing NF-kappaB activation prevents E-induced iNOS upregulation &its consequences. Significance: Showing that ALD requires a leaky gut, &that NO &cytoskeletal pathways are involved, could 1) identify drinkers at risk for LD (sugar test);2) lead to therapies to prevent LD in those drinkers unable to abstain.